SCIDA (T-B-NK+ severe combined immunodeficiency of Athabascan-speaking Native Americans) is a primary immune deficiency of humans resulting from absence of Artemis protein, required for nonhomologous end-joining in V(D)J gene rearrangement. Currently, the only effective therapy for SCIDA is allogeneic bone marrow transplant, for which there is a high risk of morbidity and mortality. Recently, the laboratory of collaborating investigator Dr. Mort Cowan, generated a mouse model of SCIDA by homologous disruption of the murine Scida gene in embryonic stem cells. Here we propose to use this Scida+ mouse model as a test system to evaluate two different gene therapy approaches for SCIDA. In Aim 1, lentiviral vectors based on HIV-1 will be designed for optimized expression of the Scida cDNA sequence in lymphoid cell populations. These vectors will first be tested in Scida -/-mouse embryo fibroblasts (MEF's) for transduction and restoration of non-homologous end-joining activity. These vectors will then be used to introduce the Scida gene into Scida -/- marrow cells, evaluating recipient animals for the presence and selective outgrowth of positively transduced cell populations as well as for correction of immunodefiency. In Aim 2, nonviral vectors based on the Sleeping Beauty transposon system will be designed for introduction and optimized expression of the Scida coding sequence, first testing for the feasibility of transposition in Scida -/- mouse embryo fibroblasts. These vectors will then be electroporated into Scida -/- marrow cells (unenriched marrow, marrow enriched for stem cells, and marrow enriched for common lymphoid progenitor cells) with subsequent evaluation for engraftment, selective outgrowth and correction of immunodeficiency after transplantation into Scida -/- recipients. Results from these studies are anticipated to provide insight into the conditions under which introduction of the SCIDA gene into appropriate lympho-hematopoietic progenitors can lead to correction of immunodeficiency in scida -/- animals, with the ultimate goal of applying such gene transfer approaches to the treatment of SCIDA in humans.